Rat angiotensinogen and des(angiotensin I)angiotensinogen: purification, characterization, and partial sequencing.
Rat angiotensinogen was completely purified by a six-step procedure including (1) ammonium sulfate precipitation, (2) affinity chromatography on Affi-gel blue, (3) chromatography on DEAE-Sephacel, (4) chromatography on hydroxylapatite, (5) chromatography on Ultrogel AcA 54, and (6) isoelectric focusing. Two peaks of pure angiotensinogen were obtained, distinguishable by their isoelectric points (4.55 and 4.75). Both contained 23 microgram of angiotensin I/mg of protein. Sodium dodecyl sulfate--polyacrylamide gel electrophoresis of the peak with pI = 4.55 revealed two protein bands (respectively Mr 57000 and 59000) and a single protein band (Mr 57000) for the peak with pI = 4.75. The molecular weight of the latter homogeneous form, as determined by sedimentation equilibrium, was 55000. Only one immunoprecipitin line was observed when antiserum reacted with the heterogeneous angiotensinogen in Ouchterlony gels. The first 17 amino acids of the N-terminal region of the angiotensinogen with pI = 4.75 are reported. The amino acids in positions 10 and 11 which correspond to the renin cleavage site are leucyl-leucyl. The des(angiotensin I)angiotensinogen obtained after hydrolysis of angiotensinogen with pure mouse submaxillary gland renin was found to consist of a single protein band with an Mr of 56000 as revealed by sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Only one N-terminal residue (leucyl) was obtained for this des(angiotensin I)angiotensinogen. These findings establish that renin only cleaves angiotensinogen at a single site.
D000595:Amino Acid Sequence; D000803:Angiotensin I / Q000031:analogs & derivatives*; D000808:Angiotensinogen / Q000097:blood / Q000302:isolation & purification*; D000809:Angiotensins / Q000302:isolation & purification*; D000818:Animals; D006736:Horses; D006801:Humans; D005779:Immunodiffusion; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D010446:Peptide Fragments / Q000097:blood / Q000302:isolation & purification*; D051381:Rats; D011919:Rats, Inbred Strains; D012083:Renin / Q000378:metabolism; D013045:Species Specificity
D000809:Angiotensins; D010446:Peptide Fragments; C018899:des-angiotensin I renin substrate; D000808:Angiotensinogen; D000803:Angiotensin I; D012083:Renin